Critical Roles of the Pentose Phosphate Pathway and GLN3 in Isobutanol-Specific Tolerance in Yeast
Critical Roles of the Pentose Phosphate Pathway and GLN3 in Isobutanol-Specific Tolerance in Yeast
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DOI：
10.1016/j.cels.2019.10.006
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发表时间：
2019-12-18
期刊：
影响因子：
9.3
通讯作者：
Avalos, Jose L.
中科院分区：
文献类型：
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作者：
Kuroda, Kouichi;Hammer, Sarah K.;Avalos, Jose L.
支链醇是有吸引力的先进生物燃料;然而，它们的细胞毒性是工程微生物以高滴度生产它们的障碍。我们对酿酒酵母基因缺失文库进行了全基因组筛选，以鉴定参与异丁醇特异性耐受性的细胞系统。戊糖磷酸途径基因GND 1或ZWF 1的缺失导致对异丁醇的超敏反应，但对乙醇不敏感。相比之下，GLN 3的缺失增加了酵母对支链醇的特异性耐受性。转录组学分析表明，异丁醇诱导氮饥饿反应，通过GLN 3和GCN 4，上调氨基酸的生物合成和氮清除，同时下调糖酵解，细胞壁生物合成和膜脂质的生物合成。通过删除GLN 3来破坏这种反应足以增强耐受性，并使工程菌株的异丁醇产量提高4.9倍。这项研究说明了耐受压力的适应机制如何导致化学生产微生物发酵的毒性，以及遗传干预如何通过逃避此类机制来提高产量。
Branched-chain alcohols are attractive advanced biofuels; however, their cellular toxicity is an obstacle to engineering microbes to produce them at high titers. We performed genome- wide screens on the Saccharomyces cerevisiae gene deletion library to identify cell systems involved in isobutanol-specific tolerance. Deletion of pentose phosphate pathway genes GND1 or ZWF1 causes hypersensitivity to isobutanol but not to ethanol. By contrast, deletion of GLN3 increases yeast tolerance specifically to branched-chain alcohols. Transcriptomic analyses revealed that isobutanol induces a nitrogen starvation response via GLN3 and GCN4, upregulating amino acid biosynthesis and nitrogen scavenging while downregulating glycolysis, cell wall biogenesis, and membrane lipid biosynthesis. Disruption of this response by deleting GLN3 is enough to enhance tolerance and boost isobutanol production 4.9-fold in engineered strains. This study illustrates how adaptive mechanisms to tolerate stress can lead to toxicity in microbial fermentations for chemical production and how genetic interventions can boost production by evading such mechanisms.